Synergic Induction of Autophagic Cell Death in Anaplastic Thyroid Carcinoma

Abstract

Anaplastic thyroid carcinoma (ATC) has poor prognosis, high mortality rate and lack of effective therapy. A synergic combination of PD-L1 antibody together with cell death promoting substances like deacetylase inhibitors (DACi) and multi-kinase inhibitors (MKI) could sensitize ATC cells and promote decay by autophagic cell death. The PD-L1-inhibitor atezolizumab synergized with panobinostat (DACi) and sorafenib (MKI) leading to significant reduction of the viability, measured by real time luminescence, of three different patient-derived primary ATC cells, of C643 cells and follicular epithelial thyroid cells too. Solo administration of these compounds caused a significant over-expression of autophagy transcripts; meanwhile autophagy proteins were almost not detectable after the single administration of panobinostat, thus supporting a massive autophagy degradation process. Instead, the administration of atezolizumab caused an accumulation of autophagy proteins and the cleavage of the active caspases 8 and 3. Interestingly, only panobinostat and atezolizumab were able to exacerbate the autophagy process by increasing the synthesis, the maturation and final fusion with the lysosomes of the autophagosome vesicles. Despite ATC cells could be sensitized by atezolizumab via the cleavage of the caspases, no reduction of cell proliferation or promotion of cell death was observed. The apoptosis assay evidenced the ability of panobinostat alone and in combination with atezolizumab to induce the phosphatidil serine exposure (early apoptosis) and further the secondary necrosis. Instead, sorafenib was only able to cause necrosis. The increase of caspases activity induced by atezolizumab, the apoptosis and autophagy processes promoted by panobinostat synergize thus promoting cell death in well-established and primary anaplastic thyroid cancer cells. The combined therapy could represent a future clinical application for the treatment of such lethal and untreatable solid cancer.

Keywords:

• Autophagy
• cell death
• PD-L1
• atezolizumab
• histone deacetylase inhibitor
• tyrosine kinase inhibitor
• anaplastic thyroid cancer

Acknowledgements

We are thankful to Michael Wanzel and Thorsten Stiewe (Institute for Molecular Oncology, Philipps University Marburg, Germany) for supporting our study by allowing us to perform the autophagy assay in the Incucyte device. We are thankful to Tamotsu Yoshimori for providing the GFP-RFP-tag- LC3B plasmid.

Author contributions

Conceptualization, Sabine Wächter, Katharina Holzer, Detlef Bartsch and Pietro Di Fazio; Data curation, Franziska Knauff, Silvia Roth, Corinna Keber and Pietro Di Fazio; Formal analysis, Franziska Knauff, Corinna Keber and Pietro Di Fazio; Funding acquisition, Sabine Wächter; Investigation, Franziska Knauff, Silvia Roth, Corinna Keber and Pietro Di Fazio; Methodology, Silvia Roth, Corinna Keber and Pietro Di Fazio; Project administration, Sabine Wächter and Pietro Di Fazio; Resources, Sabine Wächter, Katharina Holzer, Jerena Manoharan, Elisabeth Maurer and Detlef Bartsch; Validation, Sabine Wächter and Pietro Di Fazio; Visualization, Franziska Knauff, Detlef Bartsch and Pietro Di Fazio; Writing—original draft, Sabine Wächter and Pietro Di Fazio; Writing—review & editing, Katharina Holzer, Jerena Manoharan, Elisabeth Maurer, Detlef Bartsch and Pietro Di Fazio.

Declaration of interest

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the article.

Additional information

Funding

This research was funded by Anneliese Pohl-Habilitationsförderung to S.W.